Forskolin-stimulated cAMP accumulation

Yamaguchi I, Harmon SK, Todd RD, O Malley KL (1997) The rat D4 dopamine receptor couples to cone transducin (Galphat2) to inhibit forskolin-stimulated cAMP accumulation. J Biol Chem 272 16599-16602. 

Bld-HrTH administration to B. discoidalis in vivo or to isolated fat body fails to stimulate either fat body cAMP levels or adenylate cyclase activity and supports the previous findings (25). Nevertheless, for B. discoidalis, fat body phosphorylase activity is elevated and trehalose levels increase both in vivo and in vitro, and calcium is essential in vitro in addition to Bld-HrTH. No stimulation of trehalose synthesis is noted with agents that elevate adenylate cyclase, such as forskolin, or by inhibitors of phosphodiesterase such as theophylline or isobutylmethylxanthine (IBMX). Additions of cAMP, dibutyryl cAMP or 8-bromo-cAMP are not stimulatory to trehalose synthesis either in vivo or in vitro. This same result was observed for P. americana in that neither cAMP nor dbcAMP stimulated trehalose production by fat body in vitro, and xanthine inhibitors of phosphodiesterase that should cause accumulation of intracellular cAMP were inhibitory, except for isobutylmethylxanthine (IBMX) which was stimulatory for unknown reasons (26). We have not observed a stimulatory effect by IBMX with B. discoidalis fat body in vitro. 

The ability of selected agonists to stimulate phospho-inositide accumulation (for M, Mj, and Mj receptors) or to inhibit forskolin-stimulated accumulation of cAMP (for Mj and M4 receptors) was determined in CHO cells transfected with (Schwarz, 1993). 

Fig. 3. Functional study with the split chimeric Mj-trunc/Mj-tail receptor. (a) Carbachol fails to induce inositol monophosphate (IP ) accumulation in COS-7 cells transfected with the split chimeric Mj-trunc/M 3-tail receptor. Responses are expressed as percentages of increases in IP, above basal levels determined in the absence of carbachol. (b) Carbachol fails to inhibit the forskolin stimulated cAMP accumulation in cells transfected with the split chimeric M2-trunc/M3-tail receptor. The effect of carbachol is represented as a percentage of the maximal stimulated cAMP content by 100 p,M forskolin. Note that the split Mj-trunc/Mj-tail and Mj-trunc/Mj-tail receptors are as potent as the wild type M3 and Mj receptors, (c) Effect of CTPyS on carbachol inhibition curve in COS-7 cells transfected with the split chimeric M2-trunc/M3-tail receptor.

In functional experiments we studied whether or not the split chimeric M2-trunc/M 3-tail receptor retains in part the functional activity of the wild type M2 and M3 receptors. Carbachol stimulation of COS-7 cells co-transfected with the split chimeric M2-trunc/M 3-tail receptor neither inhibited forskolin-stimulated cAMP accumulation nor induced inositol monophosphate accumulation (Fig. 3a and b). It is important to note that carbachol stimulation of COS-7 cells co-transfected with fragments originating from the same receptor, M2-trunc/M2-tail and M3-trunc/M3-tail resulted respectively in the inhibition of adenylate... 

Note intrinsic Activity compared to the full DA agonist quinpirole inhibition of forskolin-stimulated cAMP accumulation in GH4C1 cells transfected with the DA D2 receptor. 

When the human PP1/Y4 receptor was stably expressed in CHO cells, it inhibited forskolin-stimulated cAMP accumulation by 50% in response to human PP at 100 nM with EC50 at 7 nM (Lundell et al., 1995). In mouse fibroblasts (thymidine kinase... 

Several effects of forskolin on B-lymphocytes, the cells of the immune system responsible for the production of immunoglobulins, have further been reported. This diterpene was found to inhibit cellular proliferation of B cells stimulated either by antibodies to surface immunoglobulins (anti-mu), and an antibody to CD20 antigen or 12-O-tetradecanoyl phorbol 13-acetate [219]. There was also a clear inhibition of G1 entry and DNA synthesis, and forskolin maintained its inhibitory effect even when added later after anti-mu stimulation. Additionally, no differences were found in the inhibitory effect of forskolin on neoplastic B cells, as compared to the responses of normal cells. Growth inhibition associated with an accumulation of cells in G1 was later found when cells of the B-lymphoid precursor cell line Reh were incubated with forskolin [220]. In that study, a delay of cells in G2/M prior to G1 arrest was observed, suggesting that important restriction points located in the G1 and G2 phases of the cell cycle may be controlled by forskolin (due to cAMP levels elevation). In a subsequent study [221], it was found that the arrest of Reh cells was accompanied by rapid dephosphorylation of retinoblastoma protein, which was suggested to be a prerequisite for the forskolin mediated arrest of these cells in Gl. 

There is general agreement that GRF uses cAMP as a second messenger system. It has been known for many years that cAMP derivatives stimulate GH in pituitary in in vitro preparations [50,51], Forskolin and cholera toxin have a similar effect [52], GRF stimulates the accumulation of intracellular cAMP in cultured pituitary cells as early as 1 min after its addition [53]. In further support of the messenger role of cAMP is the finding that the addition of various agents increasing intracellular cAMP levels to a maximally effective dose of GRF does not further increase... 

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